单核细胞上TLR4与慢性牙周炎的相关性研究

发布时间：2018-03-25 06:03

本文选题：慢性牙周炎　切入点：单核细胞　出处：《锦州医科大学》2016年硕士论文

【摘要】：目的探究单核细胞上Toll样受体4(Toll-like Receptor 4,TLR4)与人慢性牙周炎(Chronic Periodontitis,CP)发病的关系。方法于辽宁医学院附属第二医院,随机选取被诊断为CP的患者15例,牙周健康者(Healthy Control,HC)10例,取其外周血,应用流式细胞术(Flow Cytometry,FCM)检测CD14+单核细胞上TLR4及Interleukin(IL)-6的表达变化;酶联免疫吸附测定法(Enzyme-linked Immunosorbent Assay,ELISA)检测血浆中IL-6的水平变化。初步探究单核细胞上TLR4在CP发病机制中的作用。细胞实验进一步验证外周血实验结果,即:培养THP-1细胞后,分别给予0.9%氯化钠注射液(NS)、脂多糖(LPS)、TLR4特异性阻断剂(Anti-TLR4)及Anti-TLR4+LPS处理,6h后,分别采用FCM、实时荧光定量PCR检测TLR4的表达,用ELISA技术检测细胞培养上清中IL-6的水平变化。比较NS、LPS及Anti-TLR4组TLR4及IL-6的表达变化,以验证外周血实验结果;比较Anti-TLR4与Anti-TLR4+LPS组TLR4和IL-6的表达变化,探究TLR4介导的信号通路是否可以成为CP的治疗靶点。结果1.CP组外周血单核细胞上TLR4与IL-6的平均荧光强度均高于HC组(P0.001)。2.CP组血浆中IL-6的水平亦高于HC组(P0.001)。3.CP患者单核细胞内IL-6的表达与TLR4的表达呈正相关(P0.01),血浆中IL-6的水平与单核细胞内IL-6的表达亦呈正相关(P0.01)。4.LPS组THP-1细胞上TLR4的在蛋白与m RNA水平的表达及细胞上清中IL-6的水平均高于NS组(P=0.019,P0.001,P0.001);特异性阻断TLR4后,与LPS组相比,TLR4的表达及细胞上清中IL-6的水平均降低(P0.001)。5.Anti-TLR4与Anti-TLR4+LPS两组细胞上TLR4的表达和细胞上清中IL-6的水平无差异。结论G-来源的LPS可引起单核细胞上TLR4的表达升高,并通过TLR4而激活单核细胞,使IL-6等致炎因子水平上调,参与慢性牙周炎的病理形成。TLR4识别LPS信号通路可成为慢性牙周炎的治疗靶点。
[Abstract]:Objective to investigate the relationship between 4(Toll-like Receptor 4 (TLR4) and chronic periodontitis in human chronic periodontitis. Methods 15 patients with CP and 10 patients with periodontal health were randomly selected from the second affiliated Hospital of Liaoning Medical College. Flow Cytometry (FCM) was used to detect the expression of TLR4 and Interleukin(IL)-6 in CD14 monocytes. Enzyme linked immunosorbent assay (Elisa) was used to detect the level of IL-6 in plasma. The role of TLR4 on monocytes in the pathogenesis of CP was preliminarily explored. After being treated with 0.9% sodium chloride injection (NSN), lipopolysaccharide (LPSS-TLR4) specific blocker (Anti-TLR4) and Anti-TLR4 LPS for 6 h, the expression of TLR4 was detected by real-time fluorescence quantitative PCR. ELISA technique was used to detect the level of IL-6 in the supernatant of cell culture, to compare the expression of TLR4 and IL-6 in NSN LPS group and Anti-TLR4 group in order to verify the results of peripheral blood experiment, to compare the changes of TLR4 and IL-6 expression between Anti-TLR4 group and Anti-TLR4 LPS group, and to compare the expression of TLR4 and IL-6 between Anti-TLR4 group and Anti-TLR4 LPS group. Results the average fluorescence intensity of TLR4 and IL-6 in peripheral blood monocytes in 1.CP group was higher than that in HC group P0.001n.2.2. the level of IL-6 in plasma of 1.CP group was higher than that of HC group P0.001n.3.3. The expression of IL-6 was positively correlated with the expression of TLR4, and the level of IL-6 in plasma was positively correlated with the expression of IL-6 in monocytes. 4. The expression of TLR4 in THP-1 cells and the level of IL-6 in the supernatant of THP-1 cells were higher than those in LPS-treated THP-1 cells. In NS group, P0. 019, P0. 001, P0. 001, and after blocking TLR4, Compared with the LPS group, the expression of TLR4 and the level of IL-6 in the supernatant of TLR4 were significantly lower than those in the Anti-TLR4 LPS group. 5. The expression of TLR4 and the level of IL-6 in the supernatant of the two groups were not significantly different from those in the Anti-TLR4 LPS group. Conclusion G- derived LPS can induce the increase of TLR4 expression in monocytes. Monocytes were activated by TLR4, and the level of IL-6 and other inflammatory factors were up-regulated. TLR4 was involved in the pathogenesis of chronic periodontitis. TLR4 recognition of LPS signal pathway could become the therapeutic target of chronic periodontitis.
【学位授予单位】：锦州医科大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R781.4

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